The present invention pertains to methods and apparatus for imaging at resolutions below that corresponding to the wavelength of illuminating radiation.
The ability to resolve smaller and smaller features using microscopy has resulted in enormous gains in almost all branches of science and technology. Conventional optical light microscopy has been used for centuries, and can resolve features down to the Abbe-Rayleigh diffraction-limit, which is one-half the wavelength of the illumination and thus, for imaging at visible wavelengths, is typically a length scale from 150-350 nm. Comparing this resolution to atomic scales, the diameter of a hydrogen atom is 0.053 nm and the lattice spacing a silicon crystal is 0.51 nm, so there are orders of magnitude of size between conventional microscope resolution and atomic resolution.
While the conventional microscope has been sufficient to image entire biological cells and some of their constituents, continuing improvements in biology and nanotechnology have made attaining sub-optical-wavelength resolution more important. To this end, a number of microscopy methods using radiation other than visible light have been developed. Scanning Electron Microscopy (SEM) and Transmission Electron Microscopy (TEM) use an electron beam to probe surface features, and have a much better resolution limited only by the much smaller wavelength of the electron. Unfortunately these methods require the sample to be placed in a vacuum, which is harmful for many biological samples, and subjects the sample to ionizing and heating radiation often damaging the sample.
Another method, Scanning Tunneling Microscopy, (STM) operates by scanning an atomically sharp conducting probe over the surface, and measures the electric current produced by quantum mechanical tunneling at each point on the surface. Atomic Force Microscopy (AFM) scans a tip attached to a cantilever over the surface and measures the atomic forces exerted on the tip by measuring the deflection of the cantilever. While both STM and AFM have produced excellent atomic-scale images, they are slow because they require serial scanning of the surface, moreover, near-contact of the probe and surface can result in surface alteration or tip damage. In addition, the resolution achievable by these methods is dependent on the exact shape of the tip, which must be made highly uniformly to produce a repeatable image.
These disadvantages have motivated the creation of methods using visible light that can resolve features below the Abbe-Rayleigh limit. The Abbe-Rayleigh limit exists because when light is scattered off of surfaces it is encoded into both homogeneous and evanescent waves. Homogeneous waves (such as plane waves) can propagate through empty space freely, and therefore can be sensed at a location remote from the scatterer. Unfortunately, these waves contain only information corresponding to features greater than one-half the wavelength of light on the surface. The evanescent waves, which correspond to the features less than one-half the wavelength, do not propagate away from the scatterer more than a few wavelengths at most. The magnitude of the evanescent waves decays exponentially very quickly away from the scatterer, so that even a few wavelengths from the scatterer the evanescent waves are not directly detectable. Because of this, a conventional lens, which is typically many wavelengths away from the scatterer, does not receive or relay the evanescent waves. This is what limits conventional microscopy to the Abbe-Rayleigh limit. However, if a probe is placed less than a wavelength from a scatterer, the probe can scatter a portion of the evanescent waves in the vicinity of the scatterer into homogeneous waves, which then can be detected in the far field. This principle is the basis for near-field microscopy instruments.
The term “near-field,” as used herein and in any appended claims, shall have the meaning of a regime in which evanescent components of a scattering wavefunction are significant.
The most widely used methods of near-field microscopy are now discussed with reference to FIGS. 1(a) and 1(b). The Near-Field Scanning Optical Microscope (NSOM), depicted schematically in FIG. 1A and designated generally by numeral 10, places a probe 12 in the near field of a sample 8, typically between 10-200 nm from the sample 14. This probe is scanned serially over the surface, which is illuminated by uniform laser light. A typical configuration is to place the sample (equivalently referred to, herein, as the “object”) on a transparent triangular prism. The laser is incident on a surface under the sample so that it reflects off of the interior surface of the prism by total internal reflection. In the absence of the sample, the beam only extends a small fraction of a wavelength outside of the prism because the wave is evanescent. The sample perturbs the surface of the prism and scatters light from the evanescent field, some of which is collected by the probe. The light captured by the probe is guided by an optical fiber to a photodetector, where the collected photons are counted.
At each point on the surface, the photons scattered by the probe from the optical waves near the surface are collected and counted to produce an image. Typically the probe consists of a silica optical fiber, which has been heated and drawn to form a pencil-like tip on the end. The end of the fiber is then coated with a thin layer of metallic conductor. The tip of the pencil at the end of the fiber is then removed to form a tiny aperture surrounded by metal, often 10-20 nm in diameter, but open at the end. When this tip is placed in the near field of a scatterer, some of the field near the aperture at the end of the fiber is coupled into the optical fiber, where it travels up the fiber to be collected and counted at the distal end. The size of the aperture and the proximity to the sample determine the achievable resolution of the NSOM instrument.
The probe (or sometimes the sample) is translated in three dimensions with nanometer precision, typically with piezoelectric transducers. Often this is achieved in practice by integrating the NSOM instrument with an Atomic Force Microscope (AFM). The AFM measures the force between the surface 8 and a sharp probe at the end of a cantilever. By maintaining a constant force between the surface and the probe, the shape of the surface can be mapped out by determining the positions of the probe while a particular force magnitude is maintained between the surface and probe. To integrate the NSOM into the AFM, the tip of the probe is made of a material that will scatter light from the near-field of the sample. This light is collected by a lens and imaged onto a photodetector.
Apertureless NSOM, depicted schematically in FIG. 1(b), uses a sharp tip metal probe 18 supported by cantilever 17 to scatter light from the near-field to the far field, where it is collected by a lens 19. This method likewise employs serial scanning techniques, and has a resolution limited by the tip size. The probe (or sometimes the sample) is translated in three dimensions with nanometer precision, typically with piezoelectric transducers. Often this is achieved in practice by integrating the NSOM instrument with an Atomic Force Microscope (AFM). The AFM measures the force between the surface and a sharp probe at the end of a cantilever. By maintaining a constant force between the surface and the probe, the shape of the surface can be mapped out by determining the positions of the probe while a particular force magnitude is maintained between the surface and probe. To integrate the NSOM into the AFM, the tip of the probe is made of a material that will scatter light from the near-field of the sample. This light is collected by a lens 19 and imaged onto a photodetector.
While the foregoing near-field methods have been able to achieve high-resolution images (of approximately 20 nm resolution or better), they suffer from other disadvantages. First, the tip must be serially scanned over the surface. This means that the acquisition occurs only one point at a time and is very slow. Furthermore, while the entire surface is being illuminated with light, only the region near the tip provides signal at any given instant. This means that much of the available optical signal is not collected but could provide improved signal quality. Finally, in order to improve the resolution of the instrument, the aperture at the end of the probe must be made smaller. Unfortunately, this means that the probe also collects less light, further decreasing available signal. All three of these deficiencies are addressed in accordance with embodiments of the invention by gathering data from the entire scattering surface simultaneously while not requiring small apertures to achieve high resolution.